Other workers have shown that the glucose-6-phosphate dehydrogenase (G6PDH) in "old" animals and in cultured fibroblasts of humans with accelerated aging diseases (progeria, Werner's syndrome) is less active and is more sensitive to heat inactivation than is the G6PDH in young animals or in cultured cells from normal individuals. Results of the present study show that the exposure of pure G6PDH from bovine adrenals to a mixed-function oxidation system comprised of ascorbate, Fe(II) and O2 leads to inactivation of the enzyme and to the oxidation of some amino acid residues to carbonyl derivatives. The substrates NADPH and glucose-6-P protect the enzyme from this inactivation. The residual activity that remains after only 50% of the G6PDH has been inactivated is much less stable to heat inactivation than is a control sample that had been incubated in the absence of ascorbate and Fe(II). The results demonstrate that previously observed age-related changes in G6PDH activity might be caused by MFO mediated oxygen radical damage. It was incidentally discovered that G6PDH is a "cold-sensitive" enzyme. At 4 C, the enzyme dissociates to less active subunits. This inactivation can be reversed by raising the temperature to 37 C. The cold inactivation is prevented by the presence of NADPH but is accelerated by glucose-6-P. The amino acid residue in enzymes that is oxidized to a carbonyl derivative by MFO systems has not been identified. Studies with amino acid homopolymers show that a MFO system comprised of ascorbate, Fe(II), EDTA and O2 catalyze the generation of carbonyl groups in polylysine, polyhistidine, polyarginine, polyproline, and polyserine. Thus, any one of several amino acids can be a target for MFO catalyzed oxidation.